The proposed research addresses the possibility that endogenous ligands for the cannabinoid receptor play a significant role within the neural circuits that modulate pain sensitivity. The molecular basis for this hypothesis is the discovery and cloning of a G-protein coupled cannabinoid receptor and the identification of the putative endogenous cannabinoid anandamide. This laboratory showed that synthetic cannabinoids produce analgesia when injected in the dorsal periaquaductal gray (PAG), and electrical stimulation of this area produces analgesia that can be blocked by a cannabinoid antagonist suggesting that the analgesia is mediated by an endogenous cannabinoid. A recently developed method indicated the endogenous cannabinoid anandamide is released in the PAG under these stimulation conditions and upon noxious stimulation of the hindpaws. The proposed experiments further examine the possibility that the endogenous cannabinoid anandamide plays a role in pain modulation by examining its release in the periaquaductal gray (PAG), a brain region known to be part of an analgesic circuit. Release will be measured using microdialysis with high performance liquid chromatography/atmospheric pressure chemical ionization mass spectrometry. Basal release, and release stimulated by noxious stimulation or electrical depolarization are examined. The experiments will determine the peripheral fiber types and the spinal cord neurons and neural pathways that induce the release of anandamide.